Cajaninstilbene acid derivatives conjugated with siderophores of 3-hydroxypyridin-4(1H)-ones as novel antibacterial agents against Gram-negative bacteria based on the Trojan horse strategy.

The low permeability of the outer membrane of Gram-negative bacteria is a serious obstacle to the development of new antibiotics against them. Conjugation of antibiotic with siderophore based on the "Trojan horse strategy" is a promising strategy to overcome the outer membrane obstacle. In this study, series of antibacterial agents were designed and synthesized by conjugating the 3-hydroxypyridin-4(1H)-one based siderophores with cajaninstilbene acid (CSA) derivative 4 which shows good activity against Gram-positive bacteria by targeting their cell membranes but is ineffective against Gram-negative bacteria. Compared to the inactive parent compound 4, the conjugates 45c or 45d exhibits significant improvement in activity against Gram-negative bacteria, including Escherichia coli, Klebsiella pneumoniae and especially P. aeruginosa (minimum inhibitory concentrations, MICs = 7.8-31.25 µM). The antibacterial activity of the conjugates is attributed to the CSA derivative moiety, and the action mechanism is by disruption of bacterial cell membranes. Further studies on the uptake mechanisms showed that the bacterial siderophore-dependent iron transport system was involved in the uptake of the conjugates. In addition, the conjugates 45c and 45d showed a lower cytotoxic effects in vivo and in vitro and a positive therapeutic effect in the treatment of C. elegans infected by P. aeruginosa. Overall, our work describes a new class and a promising 3-hydroxypyridin-4(1H)-one-CSA derivative conjugates for further development as antibacterial agents against Gram-negative bacteria.

Exploring the Localization of Siderophore-Mediated Cargo Delivery in Gram-Negative Bacteria Using 3-Hydroxypyridin-4(1H)-one-Fluorescein Probes.

The design of siderophore-antibiotic conjugates is a promising strategy to overcome drug resistance in negative bacteria. However, accumulating studies have shown that only those antibiotics acting on the cell wall or cell membrane multiply their antibacterial effects when coupled with siderophores, while antibiotics acting on targets in the cytoplasm of bacteria do not show an obvious enhancement of their antibacterial effects when coupled with siderophores. To explore the causes of this phenomenon, we synthesized several conjugate probes using 3-hydroxypyridin-4(1H)-ones as siderophores and replacing the antibiotic cargo with 5-carboxyfluorescein (5-FAM) or malachite green (MG) cargo. By monitoring changes in the fluorescence intensity of FAM conjugate 20 in bacteria, the translocation of the conjugate across the outer membranes of Gram-negative pathogens was confirmed. Further, the use of the fluorogen activating protein(FAP)/MG system revealed that 3-hydroxypyridin-4(1H)-one-MG conjugate 26 was ultimately distributed mainly in the periplasm rather than being translocated into the cytosol of Escherichia coli and Pseudomonas aeruginosa PAO1. Additional mechanistic studies suggested that the uptake of the conjugate involved the siderophore-dependent iron transport pathway and the 3-hydroxypyridin-4(1H)-ones siderophore receptor-dependent mechanism. Meanwhile, we demonstrated that the conjugation of 3-hydroxypyridin-4(1H)-ones to the fluorescein 5-FAM can reduce the possibility of the conjugates crossing the membrane layers of mammalian Vero cells by passive diffusion, and the advantages of the mono-3-hydroxypyridin-4(1H)-ones as a delivery vehicle in the design of conjugates compared to the tri-3-hydroxypyridin-4(1H)-ones. Overall, this work reveals the localization rules of 3-hydroxypyridin-4(1H)-ones as siderophores to deliver the cargo into Gram-negative bacteria. It provides a theoretical basis for the subsequent design of siderophore-antibiotic conjugates, especially based on 3-hydroxypyridin-4(1H)-ones as siderophores.

Design and Synthesis of 3-Hydroxy-pyridin-4(1H)-ones-Ciprofloxacin Conjugates as Dual Antibacterial and Antibiofilm Agents against Pseudomonas aeruginosa.

Pseudomonas aeruginosa infections are often complicated by the fact that it can easily form a biofilm that increases its resistance to antibiotics. Consequently, the development of novel antibacterial agents against biofilm-associated drug-resistant P. aeruginosa is urgently needed. Herein, we report a series of 3-hydroxy-pyridin-4(1H)-ones-ciprofloxacin conjugates that were designed and synthesized as dual antibacterial and antibiofilm agents against P. aeruginosa. A potential 2-substituted 3-hydroxy-1,6-dimethylpyridin-4(1H)-one-ciprofloxacin conjugate (5e) was identified and had the best minimum inhibitory concentrations of 0.86 and 0.43 µM against P. aeruginosa 27853 and PAO1 and reduced 78.3% of biofilm formation. In addition, 5e eradicates mature biofilms and kills living bacterial cells that are incorporated into the biofilm. Studies on the antibiofilm mechanism of conjugates showed that 5e interferes with iron uptake by bacteria, inhibits their motility, and reduces the production of virulence. These results demonstrate that 3-hydroxy-pyridin-4(1H)-ones-ciprofloxacin conjugates are potent in the treatment of biofilm-associated drug-resistant P. aeruginosa infections.

3-Hydroxypyridin-4(1H)-one Derivatives as pqs Quorum Sensing Inhibitors Attenuate Virulence and Reduce Antibiotic Resistance in Pseudomonas aeruginosa.

The development of quorum sensing inhibitors capable of decreasing the production of virulence factors is an effective strategy to overcome resistance in Pseudomonas aeruginosa due to the less selective pressure exerted on bacteria. In this study, a series of 3-hydroxypyridin-4(1H)-one derivatives bearing a 4-aminomethyl-1,2,3-triazole linker were designed and synthesized as antivirulence agents against P. aeruginosa. The most potent derivative 16e was identified as a selective inhibitor of the pqs system (IC50 = 3.7 µM) and its related virulence factor pyocyanin (IC50 = 2.7 µM). In addition, 16e exhibited moderate biofilm inhibition and significant inhibition of P. aeruginosa motility phenotypes with low cytotoxicity. Compound 16e showed an obvious antibacterial synergistic effect in combination with antibiotics such as ciprofloxacin and tobramycin in in vitro and in vivo Caenorhabditis elegans infection models. Overall, the excellent antivirulence properties of compound 16e make it a potential antibiotic adjuvant for the treatment of P. aeruginosa infections that may be advanced into preclinical development in the future.

3-Hydroxy-pyridin-4(1H)-ones as siderophores mediated delivery of isobavachalcone enhances antibacterial activity against pathogenic Pseudomonas aeruginosa.

The natural prenylated chalcone isobavachalcone (IBC) shows good antibacterial activity against Gram-positive bacteria but is ineffective against Gram-negative bacteria, most likely due to the outer membrane barrier of Gram-negative bacteria. The Trojan horse strategy has been shown to be an effective strategy to overcome the reduction in the permeability of the outer membrane of Gram-negative bacteria. In this study, eight different 3-hydroxy-pyridin-4(1H)-one-isobavachalcone conjugates were designed and synthesized based on the siderophore Trojan horse strategy. The conjugates exhibited 8- to 32-fold lower minimum inhibitory concentrations (MICs) and 32- to 177-fold lower half-inhibitory concentrations (IC50s) against Pseudomonas aeruginosa PAO1 as well as clinical multidrug-resistant (MDR) strains compared to the parent IBC under iron limitation. Further studies showed that the antibacterial activity of the conjugates was regulated by the bacterial iron uptake pathway under different iron concentration conditions. Studies on the antibacterial mechanism of conjugate 1b showed that it exerts antibacterial activity by disrupting cytoplasmic membrane integrity and inhibiting cell metabolism. Finally, conjugate 1b showed a lower cytotoxic effects on Vero cells than IBC and a positive therapeutic effect in the treatment of bacterial infections caused by Gram-negative bacteria PAO1. Overall, this work demonstrates that IBC can be delivered to Gram-negative bacteria when combined with 3-hydroxy-pyridin-4(1H)-ones as siderophores and provides a scientific basis for the development of effective antibacterial agents against Gram-negative bacteria.

Long-term stable, high accuracy, and visual detection platform for In-field analysis of nitrite in food based on colorimetric test paper and deep convolutional neural networks.

Nitrite is one of the most common carcinogens in daily food. Its simple, rapid, inexpensive, and in-field measurement is important for food safety, based on the requirements of the standard from Codex Alimentarius Commission and China. Using polyacrylonitrile (PAN) and thin layer silica gel (SG), p-aminophenylcyclic acid (SA) and naphthalene ethylenediamine hydrochloride (NEH), as carriers and chromogenic agents, respectively, PAN-NSS as nitrite color sensor is proposed. After fixing and protecting of SA and NEH with layer-upon-layer PAN, the validity period of the test paper can be prolonged from 7 days to more than 30 days. The reproducibility of PAN-NSS preparation is ensured by electrospinning. Combined with PAN-NSS, deep convolutional neural network (DCNN) and APP as a visual monitoring platform, which has the functions of rapid sampling, data processing and transmission, intuitive feedback, etc., and provides a fully integrated detection system for field detection.

Benefit-risk assessment of metal bioavailability in edible fungi by biomimetic whole digestive tracts with digestion, metabolism, and absorption functions.

As worldwide edible fungi, Lentinula edodes and Agaricus bisporus accumulate both essential and harmful metals. Metal bioavailability is important for metal benefit-risk assessment. A full functional model of digestive tracts (including digestion, metabolism, and absorption) is established. Under the digestive tract functions, the bioaccessible and bioavailable metals are released from edible fungi and absorbed by intestinal tract, respectively. Based on bioavailable metal contents in the intestine, safe dosage and maximum consumption are 43.52 g/d and 248.7 g/d for Agaricus bisporu, 20.59/328.9 g/d (for males/ female) and 132.9 g/d for Lentinus edodes; V, Co, Ni, Cu, Zn, Se, Cr, Cd and Pb in Agaricus bisporus and Lentinula edodes are absorbed mainly in the large intestine; Fe is mainly absorbed in small intestine; edible fungi species-specificity in metal bioavailability is observed for As and Mn, which are mainly absorbed by small and large intestine for Agaricus bisporus and Lentinus edodes, respectively; and then metal toxicity on small and large intestine is disclosed. Metal benefit-risk is assessed by the content of monolayer liposome-extracted metal in the chyme from small and large intestine, which is controlled by the gastrointestinal functions, metal and edible fungi species.

Evaluation of sensory function and recovery after replantation of fingertips at Zone I in children.

Sensory function is the most significant criterion when evaluating the prognosis of replanted fingers. Current clinical research has focused on surgical techniques and indications for finger replantation; however, few studies have focused on recovery of finger sensory function after replantation. This study retrospectively assessed data of eight patients who had undergone nine Zone I replantations of the fingertips in the First Affiliated Hospital of Sun Yat-sen University of China from July 2014 to January 2016. Variations in the extent of damage, with the residual vessels or nerves in some fingers being too short or even missing, prevented tension-free suture repair in some patients. Thus, repair of four of the nine fingertips included arteriovenous anastomosis, the remaining five undergoing arterial anastomosis during replantation of the amputated fingers. Three patients underwent nerve repair, whereas the remaining six cases did not. Fingertip replantations were successful in all eight patients. Compared with the patients without vascular anastomosis, no obvious atrophy was visible in the fingertips of patients who did undergo vascular anastomosis during replantation and their sensory function did recover. Fingertip replantation provides good sensory function and cosmetic outcomes when good artery and vein anastomoses have been created, even when digital nerves have not been repaired.

[Further study on possibility of diffusion of Oncomelania hupensis with water in river channels of the east route of South-to-North Water Diversion Project].

OBJECTIVE: To study the drifting law of floats and potential risks of Oncomelania hupensis diffusion in the water diversion rivers of the east route of South-to-North Water Diversion Project. METHODS: The O. hupensis snails in the river channels were monitored by the salvage method and snail luring method with rice straw curtains, and the diffusion possibility of snails along with water was assessed through the drift test of floats with GPS. RESULTS: In the flood seasons from 2006 to 2013, totally 8 338.0 kg of floats were salvaged, and 2 100 rice straw curtains were put into water in the Li Canal and Jinbao shipping channel, but no Oncomelania snails were found. The drift test of floats with GPS before water diversion showed that the flow velocity on water surface (northbound) was 0.45 m/s, the average drift velocity of the floats was 0.56 - 0.60 m/s, and the average drift distances each time were 999.70 - 1 995.50 m in the Gaoshui River section, while there were no obvious drift in Jinbao shipping channel section. During the water diversion period, the flow velocity on water surface (northbound) was 0.45 m/s, the average drift velocity of the floats was 0.35 - 0.41m/s, and the average drift distances each time were 1 248.06 -1 289.44 m in the Gaoshui River, while in Jinbao shipping channel section, the flow velocity on water surface was 0.28 m/s, the average drift velocity of the floats was 0.25 - 0.27 m/s, and the average drift distances each time were 477.76 - 496.38 m. The drift test showed that the floats gradually closed to the river bank as affected by water flow, wind direction and ship waves, when blocked by the reeds, water plants or other obstacles, and they would stopped and could not continue to drift without outside help. CONCLUSIONS: There are no Oncomelania snails found in the river channels of the east route of South-to-North Water Diversion Project. The drifting distance of the floating debris along with the water is restricted by the flow rate and shore environment.

[Effect of comprehensive measures on schistosomiasis control in an aquaculture zone].

OBJECTIVE: To evaluate the effect of comprehensive measures on the control of schistosomiasis in an aquaculture zone, so as to provide the evidence for formulating the schistosomiasis control strategy in special environments. METHODS: From 2003 to 2012, the comprehensive control measures including individual prevention, detection and treatment of schistosomiasis patients, environmental modification, and health education were implemented in an aquaculture zone of Luyang Lake, Jiangdu District of Yangzhou City, Jiangsu Province. The changes in human morbidity of schistosomiasis and the status of Oncomelania snails were investigated annually. RESULTS: No acute infection of Schistosoma japonicum was detected for successive 10 years, and only 5 cases of chronic infection were found in the latest 5 years in the aquaculture zone. Since 2007, no snail habitats were found. CONCLUSIONS: The comprehensive control measures are effective.

[Effect of sand buried on elimination of Oncomelania hupensis snails in marshland along Yangtze River in Jiangdu District, Jiangsu Province].

OBJECTIVE: To evaluate the effect of sand buried on the elimination of Oncomelania snails in the marshland along the Yangtze River downstream. METHODS: The areas along the Yangtze River were chosen as the investigated objects in Jiangdu District, Jiangsu Province. The datum review and field investigations were used to observe the change of snail area after the sand buried. RESULTS: There were 11 sand buried projects along the Yangtze River in Jiangdu District, Jiangsu Province. The snail areas decreased by 93.73% after the sand buried. CONCLUSION: The sand buried is an effective method of snail control in the marshland along the Yangtze River. This method can be combined with economic construction and development projects for popularization and application.

Functional implications of microRNA-215 in TGF-ß1-induced phenotypic transition of mesangial cells by targeting CTNNBIP1.

Mesangial cell (MC) phenotypic transition is crucial for the progression of diabetic nephropathy. A major stimulus mediating high glucose-induced MC phenotypic transition is TGF-ß1. Our current study focuses on microRNA-215 (miR-215) and investigates its role in TGF-ß1-mediated MC phenotypic transition. Using real-time quantitative PCR (qRT-PCR) and northern blotting, we determined that the miR-192/215 family is dramatically upregulated under diabetic conditions both in vitro and in vivo. Gain- and loss-of-function approaches demonstrated that miR-215 inhibition significantly inhibited TGF-ß1-induced mouse mesangial cell (MMC) phenotypic transition, whereas miR-215 upregulation promoted MMC phenotypic transition. Interestingly, these changes were not detected in cells that were treated with TGF-ß1 and miR-192 mimics or inhibitors. These results suggest that miR-215 participates in TGF-ß1-induced MMC phenotypic transition. Luciferase reporter assays were used to identify whether catenin-beta interacting protein 1 (CTNNBIP1) is a direct target of miR-215, which was predicted by bioinformatic analysis. Mechanistic studies revealed that CTNNBIP1 suppresses Wnt/ß-catenin signaling and that miR-215 promotes ß-catenin activation and upregulates α-SMA and fibronectin expression in TGF-ß1-treated MMCs by targeting CTNNBIP1. In addition, in vivo miR-215 silencing with a specific antagomir significantly increased CTNNBIP1 protein expression, resulting in reduced ß-catenin activity and decreased α-SMA and fibronectin expression in db/db mouse kidney glomeruli. Taken together, our findings indicate that miR-215 plays an essential role in MC phenotypic transition by regulating the CTNNBIP1/ß-catenin pathway, which is related to the pathogenesis of diabetic nephropathy.

[Technology of preventing oncomelania snail diffusion in east route of South-to-North Water Diversion Project II. Effect of sand buried and reed protection on snail control in area of water source].

OBJECTIVE: To evaluate the effect of sand buried and reed protection on Oncomelania snail control in the area of water source of the east route of South-to-North Water Diversion Project. METHODS: The Oncomelania snail eggs were counted after the snails raised seven days in the sand of different contents in the spawning period. The survival of the snails was observed when the snails were raised on the sand surface in the laboratory. The change of the densities of living snails and reed growth were observed in the area of water source. RESULTS: The snails did not lay eggs in the pure sand environment. There was a negative correlation between the number of snail eggs and the content of sand (r = -0.965, P = 0.008). The mortality rates of the snails were increasing with the increase of the time in the sand environment. The mortality rates of the snails were 96.00% and 100% when the snails were raised 3 months and 6 months around 25 degrees C respectively. The field test showed that the snails were not discovered after the sand buried, the second spring, after the flood season, and the third spring. However, the density of living snails of the control group dropped by 93.65% 2 weeks after using molluscicide, but increased by 100% and kept in 0.37 snails/0.1 m2 after the flood season and the third year spring, respectively. The reed growth was good in the second spring after the sand buried. CONCLUSIONS: The sand environment is unfavorable for laying eggs and survival of the snails. The sand buried method has the effects of snail control and reed protection. In addition, the method could also prevent the snail spread in the flood season.

[Impact of simulation operation high water level on Oncomelania hupensis natural growth in water diversion rivers of east route of South-to-North water Diversion Project].

OBJECTIVE: To verify the impact of operation high water level on Oncomelania hupensis natural growth in the water diversion rivers of the east route of South-to-North Water Diversion Project. METHODS: The data of the daily mean water level were investigated in the stilling pool of Jiangdu Pumping Station during the water diversion under the emergency anti-drought, and the water level data of the previous year served as the control. The dynamic of natural growth of O. hupensis snails was observed before and after the high water level in the beach wetlands of the stilling pool of Jiangdu Pumping Station. The water level of emergency anti-drought was as simulation operation water level of the east route of South-to-North Water Diversion Project in the stilling pool, and then the effects of operation water level on O. hupensis natural growth in the water diversion rivers of the east route of South-to-North Water Diversion Project were studied. RESULTS: The altitude of the beach wetlands was 6.4 to 7.2 meters in the stilling pool of Jiangdu Pumping Station. The water diversion was 255 days continuously and the pumped water was 6.235 billion cubic meters in the Jiangdu Pumping Station from November 2010 to July 2011. There were 216, 136 days and 39 days when daily mean water level were more than or equal to 7.2, 7.6 and 8.0 meters during the water diversion, respectively. The snail densities were 0.856 snails/0.1m2 and 0.0153 snails/0.1m2, respectively, before and after the No.3 beach wetlands flooded in the stilling pool of Jiangdu Pumping Station. The densities of living snails decreased by 98.21%, and there was no any progeny of snail. The snails were not detected in other environment of the stilling pool. CONCLUSION: The operation high water level can effectively restrain snail breeding in the water diversion rivers of the east route of South-to-North Water Diversion Project.